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Methods Enzymol. 2013;528:237-55. doi: 10.1016/B978-0-12-405881-1.00014-8.

H2O2: a chemoattractant?

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1
Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.

Abstract

H2O2 is a relatively stable, rapidly diffusing reactive oxygen species that has been recently implicated as a mediator of leukocyte recruitment to epithelial wounds and transformed cells in zebrafish. Whether H2O2 activates the innate immune response by acting as a bona fide chemoattractant, enhancing chemoattractant sensing, or triggering production of other chemoattractive ligands remains largely unclear. Here, we describe the basic experimental procedures required to study these questions. We present a detailed protocol of the zebrafish tail fin wounding assay and explain how to use it for analyzing leukocyte chemotaxis in vivo. We further outline a method for H2O2 measurement in live zebrafish larvae using the genetically encoded sensor HyPer on a wide-field and a spinning disk confocal microscope. These methods provide a basis for dissecting the role of H2O2 in leukocyte chemotaxis in a vertebrate animal.

KEYWORDS:

Chemotaxis; DUOX; H(2)O(2); HyPer; Leukocyte; Wound; Zebrafish

[Indexed for MEDLINE]

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